Project Summary/Abstract The function of the nerves system relies on appropriate neural circuit assembly and long-term maintenance of the circuit integrity. Employing integrated single-cell genomic approaches, here I propose to investigate cellular and molecular mechanisms of developmental assembly and age-induced decline of neural circuits using the Drosophila olfactory system. In the Drosophila olfactory circuit, 50 classes of olfactory receptor neurons (ORNs; each class expresses a unique olfactory receptor) connect precisely with 50 classes of olfactory projection neurons, providing a powerful model to study the neuronal wiring specificity. Single-cell profiling methods (RNA-seq and ATAC-seq) are emerging as powerful approaches to study numerous biological questions, including neurodevelopment [22-27]. Recently, I have developed the first single-cell RNA-seq protocol for Drosophila neurons (Li et al., 2017 Cell) [25]. Here, I would like to apply single-cell profiling approaches to 1) classify ORN classes, 2) identify wiring specificity genes, and 3) explore mechanisms that coordinate olfactory receptor expression and ORN wiring specificity (Aims 1&2; mainly on K99 training phase). Loss of olfactory sensitivity is among the first signs of aging and neurodegenerative diseases, such as Alzheimer's. In Drosophila, the olfactory sensitivity also significantly declines with age. Here, I propose a mechanistic study to identify cellular and molecular mechanisms for age-related olfactory decline in Drosophila. Our lab has recently found that glia cells are critical for regulating the assembly of the fly olfactory circuit [8]. Numerous studies have shown that the glia-neuron interactions are important for the brain function in many neuronal systems [9-11]. Together, I hypothesize that glial cells continue to be required for maintaining the olfactory circuit function throughout the life, and that age-associated glial dysfunction contributes to the olfactory decline in old flies. I would like to combine single-cell profiling techniques with genetic methods to test this hypothesis (Aim 3; mainly on R00 independent phase). My mentoring team consists of Dr. Liqun Luo (mentor), a world-renowned neuroscientist, and Drs. Howard Chang (co-mentor) and Stephen Quake (collaborator), two leading scientists in single-cell genomics and epigenomics, as well as Dr. Tony Wyss-Coray (consultant), an expert on aging and neurodegeneration, and Dr. Marc Freeman (consultant), an expert on Drosophila glia. With their guidance, I believe that I will gain extensive training in my transition to be an independent PI.